Recently, DNA has been evaluated as a chiral scaffold for metal complexes to construct so called ‘DNA-based hybrid catalysts’, a robust and inexpensive alternative to enzymes. The unique chiral structure of DNA allows the hybrid catalysts to catalyze various asymmetric synthesis reactions. However, most current studies used aqueous buffers as solvents in these asymmetric reactions, where substrates/products are typically suspended in the solutions. The mass transfer limitation usually requires a long reaction time. To overcome this hurdle and to advance DNA-based asymmetric catalysis, we examined a series of ionic liquids (ILs), inorganic salts, deep eutectic solvents (DES), glymes, glycols, acetonitrile and methanol as co-solvents/additives in the DNA-based asymmetric Michael addition. In general, these additives induce indistinguishable changes to the DNA B-form duplex conformation as suggested by circular dichroism (CD) spectroscopy, but impose a significant influence on the catalytic efficiency of the DNA-based hybrid catalyst. Conventional organic solvents (e.g. acetonitrile and methanol) led to poor product yields and/or low enantioselectivities. Most ILs and inorganic salts cause the deactivation of the hybrid catalyst except 0.2 M [BMIM][CF₃COO] (95.4% ee and 93% yield) and 0.2 M [BMIM]Cl (93.7% ee and 89% yield). Several other additives have also been found to improve the catalytic efficiency of the DNA-based hybrid catalyst (control reaction without additive gives >99% ee and 87% yield): 0.4 M glycerol (>99% ee and 96% yield at 5 °C or 96.2% ee and 83% yield at room temperature), 0.2 M choline chloride–glycerol (1 : 2) (92.4% ee and 90% yield at 5 °C or 94.0% ee and 88% yield at room temperature), and 0.5 M dipropylene glycol dimethyl ether (>99% ee and 87% yield at room temperature). The use of some co-solvents/additives allows the Michael addition to be performed at a higher temperature (e.g. room temperature vs. 5 °C) and a shorter reaction time (24 h vs. 3 days). In addition, we found that a brief pre-sonication (5 min) of DNA in MOPS buffer prior to the reaction could improve the performance of the DNA-based hybrid catalyst. We have also shown that this DNA-based catalysis method is suitable for a variety of different substrates and relatively large-scale reactions. In conclusion, a judicious selection of benign co-solvents/additives could improve the catalytic efficiency of DNA-based hybrid catalyst.